Camptothecin and related analogs (Table 1) are emerging as a prominent class of agents useful in the treatment of cancer. The camptothecins display a unique mechanism of action: stabilization of the covalent binding of the enzyme topoisomerase I (topo I), an intranuclear enzyme that is overexpressed in a variety of tumor lines, to DNA. This drug/enzyme/DNA complex leads to reversible, single strand nicks. According to the fork collision model, the DNA nicks are converted to irreversible and lethal double strand DNA breaks during replication.
The camptothecin class of anticancer agents have exhibited unusual reactivity in vivo, both with respect to drug hydrolysis and blood protein interactions. These factors have hindered the pharmaceutical development and clinical implementation of camptothecins. In terms of hydrolysis, each of the camptothecins shown in Table 1 contains an α-hydroxy-δ-lactone pharmacophore.
TABLE 1Clinical candidates and FDA-approved analogs in the camptothecin family of antitumor agents  AqueousSolubilityCompoundR10R1R2R3SolubleTopotecan/TPTHCH2NH(CH3)2OHH″CDK602HC2H5NHCH(CH3)2HH ″Irinotecan/CPT-11C2H5HH ″GI-147211C/GG-211H InsolubleCamptothecinHHHH″9-ACHNH2HH″9-NC/RubitecanHNO2HH″SN-38C2H5HOHH″DB-67Si(CH3)2C(CH3)3HOHH ″MDCPTHHAt physiological pH of 7 and above this functionality is reactive, readily converting to the biologically inactive “ring opened” carboxylate form. Thus, as a result of the labile α-hydroxy-δ-lactone pharmacophore, an equilibrium is established between two distinct drug species: 1) the biologically active lactone form where the lactone ring remains closed; and 2) a biologically-inactive carboxylate form generated upon the hydrolysis of the lactone ring of the parent drug.
This hydrolysis problem with camptothecin and many analogs (e.g. 9-aminocamptothecin, 9-nitrocamptothecin) is exacerbated in human blood. In human blood and tissues, the camptothecin equilibrium of active lactone form vs. inactive carboxylate form can be strongly modulated by the presence of human serum albumin (HSA). The lactone form of camptothecin binds to HSA with moderate affinity yet the carboxylate form of camptothecin binds much more tightly than the carboxylate, displaying the 150-fold enhancement in its affinity. Thus, the preferential binding of the carboxylate form to HSA drives the equilibrium to the right in favor of the carboxylate, resulting in the lactone ring hydrolyzing more rapidly and completely (than when camptothecin is in an aqueous solution without HSA).
The development of 7-silylcamptothecins (or silatecans) has resulted in the identification of agents with improved human blood stabilities and activities. Recent rational design efforts have resulted in the identification of A,B-ring modified camptothecins displaying improved human blood stabilities combined with potent anti-topoisomerase I activities. Dual 7,10-substitution (where the 10-substituent is a hydroxy group) results in camptothecins displaying vastly improved human blood stabilities. SN-38 contains this dual 7-alkyl-10-hydroxy substitution pattern and in 1994 it was shown that these structural modifications block SN-38 from associating with the high affinity camptothecin carboxylate binding pocket on HSA.
More recently the design of another dual 7,10-modified camptothecin has been described. The new agent is 7-t-butyldimethylsilyl-10-hydroxycamptothecin (DB-67). DB-67 displays markedly improved human blood stability and potent anti-topoisomerase I anticancer activity. The design of DB-67 was based upon the following two considerations: 1) dual 7,10-substitution patterns eliminate the highly specific binding of carboxylate form over lactone form by HSA; and 2) lactone stabilization is further promoted by enhanced lipophilicity or lipid bilayer partitioning. Lipophilicity promotes camptothecin drug stability by favoring lactone partitioning into blood cells, thereby protecting the active lactone forms from hydrolysis. The key α-hydroxy-δ-lactone pharmacophore in DB-67 displays superior stability in human blood when compared with FDA-approved topotecan, CPT-11, and several other clinically relevant camptothecin analogs. DB-67 displayed a t1/2 of 130 min. and a % lactone at equilibrium value of 30 in human blood; the t-butyldimethylsilyl group enhances lipophilicity and thereby promotes drug associations with blood cells. DB-67 is 25 times lipophilic than camptothecin and readily incorporates as its active lactone form into cellular and liposomal bilayers. Equally important, the dual 7-alkylsilyl and 10-hydroxy substitution in DB-67 blocks the associations of the carboxylate form of DB-67 with the high affinity carboxylate binding pocket on HSA. Together, the enhanced lipophilicity and altered HSA interactions provide DB-67 with the highest human blood stability when compared with clinically relevant camptothecins containing the conventional α-hydroxy-δ-lactone pharmacophore.
In vitro cytotoxicity assays have shown that DB-67 is of comparable potency relative to camptothecin and 10-hydroxycamptothecin, as well as the FDA approved analogs topotecan and CPT-11. In addition, cell-free cleavage assays reveal that DB-67 forms more stable topoisomerase I cleavage complexes than camptothecin or SN-38. In terms of in vitro potency, DB-67 has been shown to display activity against human glioma in a murine model. Overall, these stability and activity profiles of DB-67 indicate how rational drug design can result in new, highly lipophilic agents displaying improved pharmacological properties.
In this invention we describe novel, highly lipophilic intermediates and prodrugs of DB-67 and other silatecans.